Note: The secondary antibody is fluorescently labeled, and thus, light sensitive. The slide should be kept in the dark during the rest of the procedure.

The secondary antibody solution is added to the slide, which is incubated for 60 minutes in the dark.

The slide is washed twice, each time for 5 minutes with PBST followed by a third wash with PBS for 5 minutes in the dark.

The slide is spin dried for 10–15 seconds using a table centrifuge prior to scanning.

Prestige Antibodies is a registered trademark of Sigma- Aldrich® Biotechnology LP and Sigma-Aldrich Co.
Alexa Fluor is a registered trademark of Molecular Probes.
TWEEN is a registered trademark of Croda International PLC.